Acute injury of major epithelial organ systems (kidney, liver, lung, etc.) is collectively a principal cause of death worldwide. Regenerative medicine promises to meet these human health challenges by harnessing intrinsic cellular processes to repair or replace damaged tissues. Epithelial morphogenesis is a hard-wired, multicellular differentiation program that dynamically integrates microenvironmental cues to coordinate cell fate processes including adhesion, migration, proliferation, and polarization. Thus, epithelial morphogenesis is an instructive mode of tissue assembly, maintenance, and repair. Three-dimensional epithelial cell cultures in natural basement membrane (BM) extracts produce hollow, spherical cyst structures and have indicated that the BM provides the critical cell adhesion ligands to facilitate cell survival, stimulate proliferation, and promote polarization and lumen formation. However, the utility of natural BMs for detailed studies is generally limited by lot-to-lot variations, uncontrolled cell adhesive interactions, or growth factor contamination. The goal of this thesis was to engineer bioartificial extracellular matrices (ECM) that would support and modulate epithelial cyst morphogenesis. We have engineered hydrogels, based on a multi-arm maleimide-terminated poly (ethylene glycol) (PEG-4MAL), that present cell adhesive molecules and enzymatic degradation substrates and promote polarized epithelial cyst differentiation in vitro. To investigate the influence of matrix physical and biochemical signals on cyst morphogenesis, we independently varied the polymer weight percentage (wt%), the density of a cell adhesion ligand (RGD), and crosslink degradation rates of the hydrogels. Then, we evaluated functional outcomes including Madin-Darby canine kidney (MDCK II) epithelial cell survival, proliferation, cyst polarization, and lumen formation. We found that cell proliferation, but not cell survival, was sensitive to the polymer wt%, which is related to elastic modulus and crosslink density. This result defined a working range of PEG-4MAL concentration (3.5% - 4.5%) that promotes robust proliferation. Analysis of mature cysts indicated that 4.0% and 4.5% gels produced cysts resembling those typically grown in type I collagen gels while 3.5% gels produced cysts with higher incidence of inverted polarity and multiple lumens. Perturbation of matrix degradability using a slow-degrading crosslink peptide or matrix metalloproteinase inhibitors showed that the rate of matrix degradation exerts major influence on cyst growth in PEG-4MAL gels. We employed 4.0% PEG-4MAL hydrogels with RGD ligand density ranging over 0 – 2000 uM to discover that (1) lumen formation was eliminated in the absence of RGD, (2) extent of lumen formation increased with increasing RGD concentration, and (3) cyst polarity was inverted below a threshold of integrin binding to RGD. Together, these results show that the biochemical and physical properties of the matrix, particularly integrin binding and matrix degradability, effectively modulate establishment of apico-basal polarity and lumen phenotypes in MDCK II epithelial cyst structures. Furthermore, these studies validate PEG-4MAL hydrogels as a powerful culture platform to enable detailed investigation of matrix-directed modulation of epithelial morphogenesis.

Η αύξηση του προσδόκιμου διαβίωσης ενθαρρύνει την ευρύτερη χρήση τεχνητών εμφυτευμάτων για την αντικατάσταση ιστών και οργάνων ή την υποστήριξη των zωτικών λειτουργιών. Οι κυριότερες κατηγορίες υλικών που δύνανται να χρησιμοποιηθούν στα τεχνητά εμφυτεύματα είναι τα μέταλλα, τα κεραμικά, τα πολυμερή και τα σύνθετα υλικά και βρίσκουν εφαρμογή σε πολλούς τομείς της ιατρικής και της οδοντιατρικής. Στην παρούσα εργασία παρατίθενται οι κατηγορίες και οι ιδιότητες των υλικών, οι απαιτήσεις που πρέπει να πληρουν και οι εφαρμογές τους. Παρουσιάζονται, επίσης, μετρήσεις τραχύτητας επιφάνειας με τη μέθοδο afm σε κεφαλές από χάλυβα που μπορούν να χρησιμοποιηθούν στην ολική αρθροπλαστική ισχίου. / The use of medical implants in order to replace or maintain tissues and organs is now wider than ever before. The major categories of materials used in such applications in medicine and dentistry are metals, polymers, ceramics and composite materials. The aim of the present project is to present the properties, the requirements and the applications of biomaterials in medical implants. It is, also, measured with afm the roughness on surfaces of metallic heads suitable for hip joints replacement.

In this work, thermosensitive hydrogels having tunable thermo-mechanical properties were synthesized. Generally the thermal transition of thermosensitive hydrogels is based on either a lower critical solution temperature (LCST) or critical micelle concentration/ temperature (CMC/ CMT). The temperature dependent transition from sol to gel with large volume change may be seen in the former type of thermosensitive hydrogels and is negligible in CMC/ CMT dependent systems. The change in volume leads to exclusion of water molecules, resulting in shrinking and stiffening of system above the transition temperature. The volume change can be undesired when cells are to be incorporated in the system. The gelation in the latter case is mainly driven by micelle formation above the transition temperature and further colloidal packing of micelles around the gelation temperature. As the gelation mainly depends on concentration of polymer, such a system could undergo fast dissolution upon addition of solvent. Here, it was envisioned to realize a thermosensitive gel based on two components, one responsible for a change in mechanical properties by formation of reversible netpoints upon heating without volume change, and second component conferring degradability on demand. As first component, an ABA triblockcopolymer (here: Poly(ethylene glycol)-b-poly(propylene glycol)-b-poly(ethylene glycol) (PEPE) with thermosensitive properties, whose sol-gel transition on the molecular level is based on micellization and colloidal jamming of the formed micelles was chosen, while for the additional macromolecular component crosslinking the formed micelles biopolymers were employed. The synthesis of the hydrogels was performed in two ways, either by physical mixing of compounds showing electrostatic interactions, or by covalent coupling of the components. Biopolymers (here: the polysaccharides hyaluronic acid, chondroitin sulphate, or pectin, as well as the protein gelatin) were employed as additional macromolecular crosslinker to simultaneously incorporate an enzyme responsiveness into the systems. In order to have strong ionic/electrostatic interactions between PEPE and polysaccharides, PEPE was aminated to yield predominantly mono- or di-substituted PEPEs. The systems based on aminated PEPE physically mixed with HA showed an enhancement in the mechanical properties such as, elastic modulus (G′) and viscous modulus (G′′) and a decrease of the gelation temperature (Tgel) compared to the PEPE at same concentration. Furthermore, by varying the amount of aminated PEPE in the composition, the Tgel of the system could be tailored to 27-36 °C. The physical mixtures of HA with di-amino PEPE (HA·di-PEPE) showed higher elastic moduli G′ and stability towards dissolution compared to the physical mixtures of HA with mono-amino PEPE (HA·mono-PEPE). This indicates a strong influence of electrostatic interaction between –COOH groups of HA and –NH2 groups of PEPE. The physical properties of HA with di-amino PEPE (HA·di-PEPE) compare beneficially with the physical properties of the human vitreous body, the systems are highly transparent, and have a comparable refractive index and viscosity. Therefore,this material was tested for a potential biological application and was shown to be non-cytotoxic in eluate and direct contact tests. The materials will in the future be investigated in further studies as vitreous body substitutes. In addition, enzymatic degradation of these hydrogels was performed using hyaluronidase to specifically degrade the HA. During the degradation of these hydrogels, increase in the Tgel was observed along with decrease in the mechanical properties. The aminated PEPE were further utilised in the covalent coupling to Pectin and chondroitin sulphate by using EDC as a coupling agent. Here, it was possible to adjust the Tgel (28-33 °C) by varying the grafting density of PEPE to the biopolymer. The grafting of PEPE to Pectin enhanced the thermal stability of the hydrogel. The Pec-g-PEPE hydrogels were degradable by enzymes with slight increase in Tgel and decrease in G′ during the degradation time. The covalent coupling of aminated PEPE to HA was performed by DMTMM as a coupling agent. This method of coupling was observed to be more efficient compared to EDC mediated coupling. Moreover, the purification of the final product was performed by ultrafiltration technique, which efficiently removed the unreacted PEPE from the final product, which was not sufficiently achieved by dialysis. Interestingly, the final products of these reaction were in a gel state and showed enhancement in the mechanical properties at very low concentrations (2.5 wt%) near body temperature. In these hydrogels the resulting increase in mechanical properties was due to the combined effect of micelle packing (physical interactions) by PEPE and covalent netpoints between PEPE and HA. PEPE alone or the physical mixtures of the same components were not able to show thermosensitive behavior at concentrations below 16 wt%. These thermosensitive hydrogels also showed on demand solubilisation by enzymatic degradation. The concept of thermosensitivity was introduced to 3D architectured porous hydrogels, by covalently grafting the PEPE to gelatin and crosslinking with LDI as a crosslinker. Here, the grafted PEPE resulted in a decrease in the helix formation in gelatin chains and after fixing the gelatin chains by crosslinking, the system showed an enhancement in the mechanical properties upon heating (34-42 °C) which was reversible upon cooling. A possible explanation of the reversible changes in mechanical properties is the strong physical interactions between micelles formed by PEPE being covalently linked to gelatin. Above the transition temperature, the local properties were evaluated by AFM indentation of pore walls in which an increase in elastic modulus (E) at higher temperature (37 °C) was observed. The water uptake of these thermosensitive architectured porous hydrogels was also influenced by PEPE and temperature (25 °C and 37 °C), showing lower water up take at higher temperature and vice versa. In addition, due to the lower water uptake at high temperature, the rate of hydrolytic degradation of these systems was found to be decreased when compared to pure gelatin architectured porous hydrogels. Such temperature sensitive architectured porous hydrogels could be important for e.g. stem cell culturing, cell differentiation and guided cell migration, etc. Altogether, it was possible to demonstrate that the crosslinking of micelles by a macromolecular crosslinker increased the shear moduli, viscosity, and stability towards dissolution of CMC-based gels. This effect could be likewise be realized by covalent or non-covalent mechanisms such as, micelle interactions, physical interactions of gelatin chains and physical interactions between gelatin chains and micelles. Moreover, the covalent grafting of PEPE will create additional net-points which also influence the mechanical properties of thermosensitive architectured porous hydrogels. Overall, the physical and chemical interactions and reversible physical interactions in such thermosensitive architectured porous hydrogels gave a control over the mechanical properties of such complex system. The hydrogels showing change of mechanical properties without a sol-gel transition or volume change are especially interesting for further study with cell proliferation and differentiation. / In der vorliegenden Arbeit wurden thermosensitive Hydrogele mit einstellbaren thermo-mechanischen Eigenschaften synthetisiert. Im Allgemeinen basiert der thermische Übergang thermosensitiver Gele auf einer niedrigsten kritischen Löslichkeitstemperatur (LCST) oder kritischer Mizellkonzentration bzw. –temperatur(CMC/ CMT). Der temperaturabhängige Übergang von Sol zu Gel mit großer Volumenänderung wurde im ersten Fall bei thermosensitiven Hydrogelen beobachtet und ist vernachlässigbar für CMC/ CMT abhängige Systeme. Die Änderung des Volumens führt zum Ausschluss von Wassermolekülen, was zum Schrumpfen und Versteifen des Systems oberhalb der Übergangstemperatur führt. Die Volumenänderung kann unerwünscht sein, wenn Zellen in das Gel eingeschlossen werden sollen. Die Gelierung im zweiten Fall beruht hauptsächlich auf der Mizellbildung oberhalb der Übergangstemperatur und weiterem kolloidalem Packen von Mizellen im Bereich der Gelierungstemperatur. Weil die Gelierung hauptsächlich von der Polymerkonzentration abhängt, kann sich das Gel bei Zugabe von Lösungsmittel leicht wieder lösen. Hier sollten thermosensitive Gele entwickelt werden, die auf zwei Komponenten beruhen. Eine Komponente sollte aus einem ABA-Triblockcopolymer mit thermosensitiven Eigenschaften bestehen, dem Poly(ethylen glycol)-b-Poly(propylenglycol)-b-Poly(ethylen glycol) (PEPE), dessen Sol-Gel-Übergang auf Mizellierung und kolloidalem Jamming der gebildeten Mizellen basiert, und einer weiteren makromolekularen Komponente, einem Biopolymer, dass die Mizellen vernetzt. Auf diese Weise sollten thermosensitive Gele realisiert werden, die keine oder nur eine kleine Volumenänderung während der Änderung der mechanischen Eigenschaften zeigen, die stabiler gegenüber Verdünnung sein sollten als klassische Hydrogele mit einem CMC-basierten Übergang und die jedoch gezielt abgebaut werden können. Die Hydrogele wurden auf zwei Arten vernetzt, entweder durch physikalisches Vermischen, bei dem die Vernetzung durch elektrostatische Wechselwirkungen erfolgte, oder durch kovalente Kopplung der beiden Komponenten. Als makromolekulare Komponente zur Vernetzung der Mizellen wurden Biopolymere (hier: die Polysaccharide Hyaluronsäure (HA), Chondroitinsulfat oder Pektin oder das Protein Gelatin) verwendet, um die Hydrogele enzymatisch abbaubar zu gestalten. Um eine starke ionische/elektrostatische Wechselwirkung zwischen dem PEPE und den Polysachariden zu erzielen, wurde PEPE aminiert, um hauptsächlich monoaminiertes bzw. diaminiertes PEPE einsetzen zu können. Die Gele, die auf der physikalischen Mischung von aminierten PEPE mit HA bestehen, zeigten im Vergleich zu PEPE bei gleicher Konzentration eine Zunahme der mechanischen Eigenschaften, wie beispielsweise dem elastischem Modulus (G′) und dem Viskositätsmodulus (G′′) bei gleichzeitiger Abnahme der Gelierungstemperatur (Tgel). Durch Variation des Gehalts an aminierten PEPE-, konnte die Tgel in einem Bereich von 27-36 °C eingestellt werden. Interessanterweise zeigten die physikalischen Mischungen mit diaminierten PEPE (HA·di-PEPE) höhere mechanische Eigenschaften (elastischer Modulus G′) und eine höhere Stabilität gegenüber Verdünnungseffekten als Mischungen mit monoaminiertem PEPE (HA·mono-PEPE). Dies zeigt den starken Einfluss der elektrostatischen Wechselwirkungen zwischen der Carboxylgruppe der HA und der Amingruppe von PEPE. Die physikalischen Eigenschaften HA·di-PEPE sind vergleichbar mit den physikalischen Eigenschaften des Glaskörpers im Auge hinsichtlich Transparenz, Brechungsindex und Viskosität. Deswegen wurde das Material hinsichtlich seiner biologischen Anwendung getestet und zeigte sich sowohl im Überstand als auch im direkten Kontakt als nichtzytotoxisch. Zukünftig wird dieses Material in weiteren Untersuchungen bezüglich seiner Eignung als Glaskörperersatz geprüft werden. Zusätzlich konnte der enzymatische Abbau der Hydrogele mit Hyaluronidase gezeigt werden, die spezifisch HA abbaut. Beim Abbau der Hydrogele stieg Tgel bei gleichzeitiger Abnahme der mechanischen Eigenschaften. Aminiertes PEPE wurde zusätzlich zur kovalenten Bindung unter Verwendung von EDC als Aktivator an Pektin und Chondroitinsulfat eingesetzt. Tgel konnte auf 28 – 33 °C eingestellt werden durch Variation der Pfropfungsdichte am Biopolymer bei gleichzeitiger Zunahme der thermischen Stabilität. Die Pec-g-PEPE Hydrogele waren enzymatisch abbaubar, was zu einer leichten Erhöhung von Tgel und zu einer Abnahme von G′ führte. Die kovalente Bindung der aminierten PEPE an HA erfolgte unter Verwendung von DMTMM als Aktivator, der sich in diesem Fall als effektiver als EDC herausstellte. Die Reinigung mittels Ultrafiltration führte zu einer deutlich besseren Aufreinigung des Produkts als mittels Dialyse. Die gegrafteten Systeme waren in Nähe der Körpertemperatur bereits im Gelstadium und zeigten eine Erhöhung der mechanischen Eigenschaften bereits bei sehr geringen Konzentrationen von 2.5Gew.%. Die höheren mechanischen Eigenschaften dieser Hydrogele erklären sich durch die Kombination der Mizellbildung (physikalische Wechselwirkung) des PEPE und der Bildung kovalenter Netzpunkte zwischen PEPE und HA. PEPE bzw. entsprechende physikalische Mischungen derselben Komponenten zeigten kein thermosensitives Verhalten bei einer Konzentration unterhalb von 16 Gew%. Diese thermosensitiven Hydrogele zeigten auch eine Löslichkeit auf Abruf durch enzymatischen Abbau. Das Konzept der Thermosensitivität wurde in 3D strukturierte, poröse Hydrogele (TArcGel)eingeführt, bei dem PEPE kovalent an Gelatin gebunden wurde und mit LDI vernetzt wurde. Das gepfropfte PEPE führte zu einer Erniedrigung der Helixbildung der Gelatinketten. Nach Fixierung der Gelatinketten durch Vernetzung zeigte das System eine Erhöhung der mechanischen Eigenschaften bei Erwärmung (34-42 °C). Dieses Phänomen war reversibel beim Abkühlen. Eine mögliche Erklärung der reversiblen Änderungen bezüglich der mechanischen Eigenschaften sind die starken physikalischen Wechselwirkungen zwischen den Mizellen des PEPE, die kovalent an Gelatin gebunden wurden. Ferner wurde durch AFM Untersuchungen festgestellt, dass bei Temperaturerhöhung (37 °C) die örtlichen elastischen Moduli (E) der Zellwände zugenommen haben. Zusätzlich wurde die Wasseraufnahme der TArcGele durch PEPE und die Temperatur (25 °C und 37 °C) beeinflusst und zeigte eine niedrigere Wasseraufnahme bei höherer Temperatur und umgekehrt. Durch die niedrigere Wasseraufnahme bei hohen Temperaturen erniedrigte sich die Geschwindigkeit des hydrolytischen Abbaus im Vergleich zu dem strukturierten Hydrogel aus reiner Gelatin. Diese temperatursensitiven ArcGele könnten bedeutsam sein für Anwendungen im Bereich Stammzellkultivierung, Zelldifferenzierung und gerichteter Zellmigration. Zusammenfassend konnte bei den thermosensitiven Hydrogelen gezeigt werden, dass die Vernetzung von Mizellen mit einem makromolekularen Vernetzer die Schermoduli, Viskosität und Löslichkeitsstabilität im Vergleich zu reinen ABATriblockcopolymeren mit CMC-Übergang erhöht. Dieser Effekt konnte durch kovalente und nichtkovalente Mechanismen, wie beispielsweise Mizell- Wechselwirkungen, physikalische Interaktionen von Gelatinketten und physikalische Interaktionen von Gelatinketten und Mizellen, realisiert werden. Das Pfropfen von PEPE führte zu zusätzlichen Netzpunkten, die die mechanischen Eigenschaften der thermosensitiven architekturisierten, porösen Hydrogele beeinflussten. Insgesamt ermöglichten die physikalischen und chemischen Bindungen und die reversiblen physikalischen Wechselwirkungen in den strukturierten, porösen Hydrogelen eine Kontrolle der mechanischen Eigenschaften in diesem sehr komplexen System. Die Hydrogele, die eine Veränderung ihrer mechanischen Eigenschaften ohne Volumenänderung oder Sol-Gel-Übergang zeigen sind besonders interessant für Untersuchungen bezüglich Zellproliferation und –differenzierung.

<p>As an increasingly prevalent chronic disease, diabetes represents one of the fastest growing health burdens to both the developed and developing world. In an effort to improve the management and treatment of diabetes, implantable sensors that continuously monitor glucose levels have become popular alternatives to patient-administered finger prick measurements of blood glucose. However, following implantation, the performance of these implants suffers from inaccurate and erratic readings that compromise their useful lives. As a result, implantable glucose sensors remain limited as a platform for the reliable management of diabetes. While the interaction between the sensor and its surrounding tissue has been posited as a culprit for erroneous in vivo sensor performance, there remains little evidence to support that theory.</p><p>This dissertation describes the effects that implant-associated tissue reactions have on implantable sensor function. Since tissue response to an implant changes over time, the overall effect of these tissue reactions is broken into two temporal phases: (1) the phase of weeks to months following implantation when a mature foreign body capsule is present around the sensor and (2) the phase of days to weeks immediately following sensor implantation when a provisional matrix of proteins and inflammatory cells envelops the sensor.</p><p>Late stage sensor responses to implantation are marked by both an attenuated sensor signal and a significant time lag relative to blood glucose readings. For this later stage of sensor response, a computational model of glucose transport through the interstitial space and foreign body capsule was derived and implemented. Utilizing physiologically relevant parameters, the model was used to mechanistically study how each constituent part of the capsular tissue could affect sensor response with respect to signal attenuation and lag. Each parameter was then analyzed using logarithmic sensitivity analysis to study the effects of different transport variables on both lag and attenuation. Results identified capsule thickness as the strongest determinant of sensor time lag, while subcutaneous vessel density and capsule porosity had the largest effects on attenuation of the sensor signal.</p><p>For the phase of early stage tissue response, human whole blood was used as a simple ex vivo experimental system. The impacts of protein accumulation at the sensor surface (biofouling effects) and cellular consumption of glucose in both the biofouling layer and in the bulk (metabolic effects) on sensor response were assessed. Medtronic Minimed SofSensor glucose sensors were incubated in whole blood, plasma diluted whole blood, and cell-free platelet poor plasma (PPP) to analyze the effects of different blood constituents on sensor function. Experimental conditions were then simulated using MATLAB to predict the relative impacts of biofouling and metabolic effects on the observed sensor responses. It was found that the physical barrier to glucose transport presented by protein biofouling did not hinder glucose movement to the sensor surface. Instead, glucose consumption by inflammatory cells was identified as the major culprit for generating poor sensor performance immediately following implantation.</p><p>Lastly, a novel, biomimetic construct was designed to mimic the in vivo 3D cellular setting around the sensor for the focused in vitro investigation of early stage effects of implantation on glucose sensor performance. Results with this construct demonstrate similar trends in sensor signal decline to the ex vivo cases described above, suggesting this construct could be used as an in vitro platform for assessing implantable glucose sensor performance.</p><p>In total, it may be concluded from this dissertation that instead of sensors "failing" in vivo, as is often reported, that different physiological factors mediate long term sensor function by altering the environment around the implant. For times immediately following implantation, sensor signals are mediated by the presence of inflammatory macrophages adhered on the surface. However, at longer times post-implantation, sensor signals are mediated not by the consumptive capacity of macrophages, but instead by the subcutaneous vessel density surrounding the sensor as well as the porosity and thickness of the foreign body capsule itself. Taken in concert, the results of this dissertation provide a temporal framework for outlining the effects of tissue response on sensor performance, hopefully informing more biocompatible glucose sensor designs in the future.</p> / Dissertation

Large infected bone defects (IBD) are very complicated to treat due to their high variability; they often require multiple procedures. Bone autografts are the gold standard for treatment but have several drawbacks, such as a need for a second surgery site, limited grafting material, and donor site morbidity. The objective of this research was to develop a moldable synthetic bone grafting material capable of releasing both antimicrobial and osteogenic drugs over a clinically relevant time course for the treatment of IBDs. Current treatment methods for large IBDs require two separate procedures to treat the bone defect and the infection. This research sought to combine these two procedures into one implantable composite bone graft substitute for the treatment IBDs. To begin, the degradation and mechanical properties of the calcium sulfate (CS) based composite material were evaluated for different compositions. Next, the controlled drug release profiles from the composite was achieved by using a shell and core system incorporating poly(lactic-co-glycolic acid) microspheres (PLGAms). The release of vancomycin from the shell began immediately and continued over the course of 6 weeks, while the release of simvastatin from the core was delayed before being released over 4 weeks. Next, an infected, critically-sized rat femoral defect model was used to test different treatment methods with and without the composite bone graft substitute. Animals treated with locally released antibiotics had survivorship rates 24% higher than those treated with systemic antibiotics, and animals that received both antibiotics and an osteogenic drug had an increased amount of bone formation at 12 weeks compared to controls. Finally, several different anti-biofilm agents were evaluated for their ability to inhibit and/or disrupt the growth of Staphylococcus aureus (S. aureus) biofilms in vitro. Lysostaphin was the only drug investigated that was able to both inhibit and disrupt S. aureus biofilms. Furthermore, lysostaphin encapsulated into PLGAms maintained its bioactivity and may be useful for future incorporation into biofilm-combating materials. The bone grafting material developed here can be used to locally deliver drugs in a temporally controlled manner to reduce the number of procedures necessary for the treatment of complex IBDs.

This thesis aimed to investigate and document the potential of applying ceramics in two specific drug delivery applications: tamper-resistant opioid formulations and transdermal enhancement protrusions. Geopolymers were developed into the matrix for a tamper-resistant formulation, aiming to protect drug substances from non-medical abuse. The synthesis conditions and excipients composition of the geopolymer-based formulation were modified in this work to facilitate a stable and extended drug delivery. Results showed that 37ºC 100% humidity for 48 hours were applicable conditions to obtain geopolymer with suitable mechanical strength and porosity. Moreover, it was found that the integration of poly(methyl acrylate) into the geopolymer-based formulation could reduce the drug release at low pH and, meanwhile, maintain the mechanical strength. Therefore, the geopolymer-based drug formulations concluded from these studies were applied in oral and transdermal delivery systems. Evidence of the tamper-resistance of geopolymer-based oral and transdermal formulations was documented and compared to the corresponding commercial opioid formulations. The results provided experimental support for the positive effects of geopolymers as drug carriers for the tamper-resistance of oral and transdermal delivery systems. Self-setting bioceramics, calcium phosphate and calcium sulfate were fabricated into transdermal enhancement protrusions in this work for the first time. Results showed that, under mild conditions, both bioceramics could form pyramid-shaped needles in the micron size. The drug release from these needles could be controlled by the bulk surface area, porosity and degradation of the bioceramics. An in vitro insertion test showed that the bioceramic microneedles had enough mechanical strength to insert into skin. Further optimization on the geometry of needles and the substrate material was also performed. The higher aspect-ratio needles with a flexible and self-swellable substrate could release most of the drug content within 4 hours and could penetrate through the stratum corneum by manual insertion. This study explored the potential application of bioceramics in transdermal enhancement protrusions and showed promising indication of their future developments.

Cell behavior is influenced to a large extent by the surrounding microenvironment. Therefore, in the body, the cellular microenvironment is highly controlled with cells growing within well-defined tissue architectures. However, traditional culture techniques allow only for the random placement of cells onto culture dishes and biomaterials. Cell micropatterning strategies aim to control the spatial localization of cells on their underlying material and in relation to other cells. Developing such strategies provides us with tools necessary to eventually fabricate the highly-controlled microenvironments found in multicellular organisms. Employing natural extracellular matrix (ECM) materials in patterning techniques can increase biocompatibility. In the future, with such technologies, we can hope to conduct novel studies in cell biology or optimize cell behavior and function towards the development of new cell-based devices and tissue engineering constructs. Herein, a novel cell patterning platform was developed on a hydrogel base of crosslinked hyaluronic acid (HA). Hydrogels are often employed in tissue engineering due to their ability to mimic the physicochemical properties of natural tissues. HA is a polymer present in all connective tissues. Cell-adhesive regions on the hydrogel were created using the RGDS peptide sequence, found within the cell-adhesive ECM protein, fibronectin. The peptide was bound to a 2-nitrobenzyl “caging group” via a photolabile bond to render the peptide light-responsive. Finally, this “caged” peptide was covalently bound to the hydrogel to form a novel HA hydrogel with a cell non-adhesive surface which could be activated with near-UV light to become adhesive. In this way, we successfully formed chemically patterned cell-adhesive regions on a HA hydrogel using light as a stimulus to form controlled cell patterns. While the majority of cell patterning strategies to date are limited to patterning one cell population and cannot be changed with time, our strategy was novel in using small, adhesive, caged peptides combined with multiple, aligned light exposure steps to allow for dynamic chemical cell patterning on a hydrogel. Multiple cell populations, even held apart from one another, were successfully patterned on the same hydrogel. Furthermore, cell patterns were deliberately modified with time to direct cell growth and/or migration on the hydrogel base.

Through whatever misfortune people have always had the need for artificial limbs. This study questions current thinking in the field of prosthetics, aiming to address the most prevalent issues affecting the amputee today, such as, fit, comfort and practicality, which have an inarguable baring on patient prostheses satisfaction. Through examination, more obscure problems encountered by users were explained, indicating how design issues and methodologies affect the present and future manufacturing process. As a result of this research a development model for the increased effectiveness of prostheses fitrnent and improvements in patient prosthetic satisfaction have been made. This has included suggestions for potential improvements in limb fitting center protocol, patient education and awareness strategies for the assessment of delivered patient needs and requirements Methods implemented during the research consisted of a comprehensive literature review of current infonnation, technical reports and patient satisfaction findings and assessment techniques. 1bis was accompanied with an investigation and evaluation of the prosthetics industry, including limb fitting, patient requirements, product/service shortfalls, rehabilitation technique and patient lifestyle. Additionally interviews and questionnaires with practitioners and users were undertaken aiding the evaluation of patient satisfaction and the identification of potential improvements in artificial limb fitment procedure. The results revealed several areas that deserved more detailed investigation, notably relating to the hypotheses, that the relationship between the levels of fit, comfort and practicality archived within the prostheses has an effect on the patient's satisfaction. Through the examination ofthis main hypothesis one of the most significant factors which emerged was the effect ofthe communication level held between the patient and prosthetist. The results of&quot;this enquiry indicated that improved patient knowledge with respect oftheir situation and an increased ability to accurately relay issues of concern to the prosthetist, facilitated the delivery of satisfactory prostheses, in turn improving its fit, comfort and practicality. In conclusion, previous conjecture as to the limited effectiveness of current prosthetics in re-establishing patients activity levels were assessed, and suggestions generated by the results of patients dissatisfaction with their limbs. These findings facilitated the realisation of new educational, protocol-based methodologies, tools and theories.

Optical biosensors are desired for the monitoring of various biochemical markers, which are relevant indicators in the treatment and diagnosis of diseases. Specifically, luminescence sensors are favorable for optical interrogation since they are highly sensitive to analyte changes and may be implemented in lifetime or intensity-based systems. In order to develop particle-based fluorescent sensors, poly(2-hydroxyethylmethacrylate) (HEMA) microspheres have been fabricated via membrane emulsification and characterized to evaluate the emulsion method and the overall process of tailoring properties to synthesize spheres of specific mean sizes. A pH-sensitive indicator seminaphthorhodafluors-4F 5-(and-6)-carboxylic acid (SNARF) was immobilized within the microspheres, and resulting sensor particles were exposed to various pH buffers to obtain a pH calibration curve based on intensity measurements. PolyHEMA microparticles were fabricated in a systematic study with measured mean sizes ranging from 8-21 um. Optical and scanning electron microscopy images revealed the formation of spherical, porous particles, which were additionally stabilized with polymer coatings. The lowest coefficient of variation value achieved was 50%, indicating the inability to produce monodisperse particles due to the dispersity of pore sizes in the membrane. SNARF was immobilized within the polyHEMA spheres, and fluorescence was observed when exposing the sensors to different pH buffers on a fluorescence microscope. Ratiometric intensity measurements for the sensor particles were obtained on a spectrofluorometer while flowing pH buffers over the immobilized spheres in a reaction chamber. The peak intensity ratio of the microparticle sensors exhibited a change in 0.9 units when decreasing the pH from 8.4 to 5.5. In the future, these pH sensing particles may be implanted alongside glucose sensing materials in order to provide valuable pH information in understanding the immune response to specific biomaterials and sensing components.

Thesis (M.SC.) - Carleton University, 2007. / Includes bibliographical references. Also available in electronic format on the Internet.